1. Field of the Invention
The present invention relates to methods for identifying pericentric inversions and incomplete chromosome exchanges as well as methods for identifying clastogenic agents based on relationships between frequencies of different chromosome aberrations.
2. Description of Related Art
Chromosome aberrations refer to rearrangements between chromosomes (interchromosomal) and/or rearrangements within a chromosome (intrachromosomal). FIGS. 1B-1F illustrate several different chromosome aberrations which a pair of chromosomes, illustrated in FIG. 1A, can undergo. The pair of chromosomes illustrated in FIG. 1A include a first chromosome 12 and a second chromosome 12'. Each chromosome includes a centromere 14, 14', and chromatids 15 including telomeric regions (16A, 16B, 16A' and 16B'). FIG. 1B illustrates a simple translocation between chromosomes 12 and 12' where a chromatid on the first chromosome is exchanged with a chromatid on the second chromosome. FIGS. 1C and 1D illustrate types of incomplete translocations. As illustrated in FIG. 1C, a portion of a chromatid from the second chromosome 12' has been transferred to the first chromosome 12 while a portion of the first chromosome has been lost. As illustrated in FIG. 1D, a portion of a chromatid from the second chromosome 12' has been transferred to the first chromosome 12 while most of the second chromosome has been lost including centromere 14. FIG. 1E is a representation of a dicentric chromosome and a chromosome fragment produced by an aberration. FIG. 1F is a representation of a reciprocal dicentric translocation.
A chromosome translocation is one particular type of chromosome aberration. A chromosome translocation refers to the movement of a portion of one chromosome to another chromosome (interchromosome rearrangement) as well as the movement of a portion of a chromosome to a different location on that chromosome (intrachromosome rearrangement). In general, chromosome translocations are characterized by the presence of a DNA sequence on a particular chromosome that is known to be native to a different chromosome or a different portion of the same chromosome.
Two chromosome translocations of particular note are dicentric and centric rings which are formed by the rejoining of the chromatids of two different chromosomes (interchromosome) and of the same chromosome (intrachromosome) respectively. Dicentric and centric rings are highly unstable and are almost always clonogenically fatal.
Symmetric translocations and pericentric inversions are more stable analogs of dicentric and centric rings. Analysis of banded chromosomes has been used to detect symmetric translocations and pericentric inversions. Banding analysis, however, is time consuming, making it ill suited for gathering statistically significant quantities of data, especially for inversions. Lucas, et al., Rapid translocation frequency analysis in humans decades after exposure to ionizing radiation, Int'l. J. Radiat. Biol., 62:53-63 (1992); Sachs, et al., Radiation Research, 133:345-350 (1993).
Incomplete reciprocal translocations, hereinafter referred to as "incompletes," are also stable translocations. Incompletes can be detected using chromosome painting where they appear as one bicolor and one solid color chromosome derivative instead of two bicolor chromosome derivatives. It has been argued that incomplete reciprocal translocations are actually "hidden reciprocal translocations" due to one translocated segment being too small to detect. Kodama, et al. Estimation of minimal size of translocated chromosome segment detectable by fluorescence in situ hybridization, Int'l J. Radiat. Biol. 71:35 (1997). This problem cannot be resolved using conventional chromosome painting or banding because of the size resolution of the small translocated piece.
Clastogenic agents are chemicals, particles, or forms of energy which cause or enhance the frequency of chromosome aberrations. Examples of different types of clastogenic agents include, but are not limited to radioactive elements such as radon, uranium and hydrogen, their decay products and high energy particles they emit. Specific examples of radioactivity related clastogenic agents include gamma rays emitted from .sup.60 Co or .sup.137 Cs, .sup.56 Fe and .sup.12 C ions, and neutrons. Clastogenic agents can also be non-energy emitting chemicals such as benzene which can cause or enhance the frequency of chromosome aberrations.
Chromosome aberrations and the clastogenic agents which cause them or enhance their frequency are of interest due to the various cancers and other genetic diseases which are associated with chromosome aberrations. For example, radon and its decay products have been determined to be the major naturally occurring radioactive carcinogens in the human environment. Brenner, D. J. , Radon Risk and Remedy 102. (W. H. Freeman. New York, 1989). It is believed that individuals receive high linear-energy-transfer (LET) charged-particle exposure from radon gases and its daughters. However, current risk estimates based on exposure to these particles are quite uncertain. A need exists for a method for determining which individuals, particularly those who later develop cancer, did, in fact, receive significant high-LET charged-particle radiation doses. A further need exists for a method for quantifying the amount of high-LET charged-particle radiation received.
Nuclear dockyard workers who develop cancer after exposure to radiation would also benefit from a method which could be used to demonstrate a causal connection between their cancer and a high-LET radiation exposure. Lucas, J. N., et al. Discrimination between leukemia-and non-leukemia-induced chromosomal abnormalities in the patient's lymphocytes. Int'l. J. Radiat. Biol., 66:185-189 (1994).
Recent measurements of A-bomb dosimetry suggest that the neutron component may dominate the equivalent dose at relevant locations. Hoshi, M., et al. Europium-152 activity induced by Hiroshima atomic bomb neutrons, Comparison with the .sup.32 P. .sup.60 Co and .sup.152 Eu activities in Dosimetry System 1986 (DS86), Health Physics, 57:831-837 (1989). In the past year, there have been arguments advanced that most of the equivalent dose to which A-bomb survivors were exposed came from densely ionizing radiation (neutrons). A need has existed since 1945 for a method for identifying and quantifying neutron exposure for A-bomb survivors.
The need for methods for detecting aberrations and for associating the occurrence of particular aberrations with particular clastogenic agents and their dosages is evidenced by discussions in the following references: Savage, J. R. K. & Holloway, M., Induction of sister-chromatic exchanges by d(42MeV)-Be neutrons in unstimulated human lymphocytes, Brit. J. Radiol. 61:231-234 (1988); Kadhim, K. A., et al., Transmission of chromosomal instability after plutonium alpha-particle irradiation, Nature 355:738-740 (1992); Brenner, D. J. and Sachs, R. K., Chromosomal "fingerprints" of prior exposure to densely ionizing radiation, Radiation Research, 140:134-142 (1994).